Cardiac arrhythmias remain a leading cause of morbidity and mortality in the US. In multiple forms of cardiac disease, arrhythmias result from disturbances in intracellular calcium (Ca) cycling, the process that normally couples electrical excitation to mechanical function. Despite significant progress, the fundamental mechanisms underlying Ca-dependent arrhythmias remain elusive, owing mainly to the complex, nonlinear nature of cardiac Ca signaling, which hinders the development of effective antiarrhythmic therapies. Based on our work in the previous funding period, we have established that Ca signaling refractoriness provides a powerful concept for understanding cardiac intracellular Ca handling at multiple biological scales (from molecular and subcellular domains to myocardial tissue and intact heart) in normal and diseased hearts. Ca signaling refractoriness refers to a state of temporary deactivation of the sarcoplasmic reticulum (SR) Ca release channels (Ryanodine receptors, RyRs) in the wake of systolic release from the sarcoplasmic reticulum (SR). This Ca signaling refractoriness is critical in maintaining stable Ca-induced Ca release (CICR) by preventing aberrant diastolic Ca release (DCR) - a cause of arrhythmias. Using this new framework, our proposed research will establish the subcellular and molecular bases of aberrant Ca release synchronization, including the refractory properties of functionally distinct Ca release units and determine how these properties are influenced by genetic and acquired intrinsic RyR defects as well as by extrinsic local Na/Ca microdomain homeostasis and SR Ca load. Our quantitative studies of aberrant Ca-excitation coupling will provide a new understanding of characteristic differences in arrhythmogenic properties of ventricular and atrial tissue including ectopic firing propensity and self-sustaining Ca/Vm oscillations. Our multiscale studies of inhibition of arrhythmogenic propensity through the targeting of both intrinsic and extrinsic mechanisms using genetic mouse models of arrhythmia and pre- clinical models of cardiomyopathy as well as preparations from failing and rejected donor human hearts. Taken together these will provide a proof-of-principle for new therapeutic strategies based on desynchronization of aberrant SR Ca release.